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A fully human anti-new coronavirus neutralizing antibody zw2g10 and its application

A fully human, monoclonal antibody technology, applied in the fields of microbiology and immunology, can solve problems such as loss of neutralizing activity

Active Publication Date: 2022-03-25
ACADEMY OF MILITARY MEDICAL SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, with the continuous evolution and mutation of SARS-CoV-2, most of the monoclonal antibodies under development have lost their neutralizing activity against one or more mutant strains. Among them, LY-CoV555 and LY-CoV016 developed by Lilly, regeneration REGN10933 developed by Yuan has lost neutralizing activity against Beta strain virus, and antibody drugs LY-CoV555-LY-CoV016 cocktail therapy, REGEN-COV cocktail therapy, and AZD7442 cocktail therapy have lost neutralizing activity against Omicron strain

Method used

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  • A fully human anti-new coronavirus neutralizing antibody zw2g10 and its application
  • A fully human anti-new coronavirus neutralizing antibody zw2g10 and its application
  • A fully human anti-new coronavirus neutralizing antibody zw2g10 and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1 Screening and preparation of human anti-SARS-CoV-2 monoclonal antibodies.

[0037] 1. Collection of blood samples.

[0038] After obtaining informed consent, 20 mL of blood samples were collected 14 days after the second immunization of the recombinant new coronavirus vaccine vaccinators for subsequent experiments.

[0039] 2. Sorting memory B cells by flow cytometry.

[0040] The collected blood samples were separated from PBMC by Ficoll density gradient centrifugation, and the process was as follows:

[0041] 1) Take fresh anticoagulated whole blood and anticoagulate with EDTA.

[0042] 2) Add the same volume of separation liquid as the blood sample into the centrifuge tube, spread the blood sample above the liquid surface of the separation liquid, and keep the interface between the two liquid surfaces clear.

[0043] 3) Trim, room temperature, horizontal rotor 800g, acceleration and deceleration speed 3, centrifuge for 30min.

[0044] 4) After centrifug...

Embodiment 2

[0118] Example 2. Analysis of the epitope recognized by antibody ZW2G10.

[0119] 1) Coating: Take the recombinant SARS-CoV-2WT S-ECD antigen, S1 antigen, RBD antigen and S2 antigen on the 96-well enzyme-linked plate the day before the experiment and dilute to a concentration of 2 μg / mL with the coating solution, and coat the enzyme label plate, 100 μL per well, and coat overnight at 4°C.

[0120] 2) Blocking: Wash 3 times with a plate washer (BIO-TEK, 405_LS) on the day of the experiment, add 100 μL of blocking solution to each well, and incubate at 37° C. for 1 hour.

[0121]3) Sample incubation: wash the plate 3 times, add 100 μL of diluent to each well except the first well, dilute the antibody to 1 μg / mL in the first well, 4-fold serial dilution, 100 μL / well, set up three replicate wells for each antibody, Incubate at 37°C for 1 h.

[0122] 4) Secondary antibody incubation: wash the plate 3 times, dilute the HPR-labeled goat anti-human IgG secondary antibody (Abcam, ab9...

Embodiment 3

[0128] Example 3: Identification of cross-binding activity of antibody ZW2G10.

[0129] The cross-binding activity between ZW2G10 and SARS-CoV-2 Variants of concern (Variants of concern) S protein was identified, the method was the same as above, and the results were as follows Figure 7 shown. ZW2G10 specifically binds to the S-ECD protein of Alpha strain, Beta strain, Gamma strain, Delta strain, and Omicron strain, and exhibits a dose-response relationship. The results showed that monoclonal antibody ZW2G10 could cross-link the S-ECD protein of Alpha strain, Beta strain, Gamma strain, Delta strain and Omicron strain.

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Abstract

The invention discloses an anti-SARS-CoV-2 fully human monoclonal antibody ZW2G10, the antibody has a unique CDR partition, and its antigen recognition epitope is located in the RBD region of the S1 protein. The antibody neutralizes the EC of the new coronavirus wild type, Alpha, Beta, Gamma, Delta and Omicron variant pseudovirus 50 They were 14.19, 14.12, 18.41, 15.59, 36.18, and 19.26 ng / mL, respectively. ZW2G10 has neutralizing activity against the current main mutant strains. The monoclonal antibody disclosed in the present invention also has the characteristics of high expression, full human origin, and good stability, is suitable for industrial production, and has great application value for responding to the outbreak caused by the new crown mutant strain.

Description

technical field [0001] The invention discloses an antibody and belongs to the fields of microbiology and immunology. Background technique [0002] The pathogen of new coronary pneumonia (COVID-19) is novel coronavirus-2 (SARS-CoV-2), and SARS-CoV-2 belongs to the β-coronavirus genus of the Coronaviridae family, and is a type of single-strand positive strand with an envelope RNA virus, its genome length is about 30kb. The first 2 / 3 of the genome is the non-structural gene ORF1a / b, which mainly encodes enzymes related to viral replication (RNA-dependent RNA polymerase, RdRp), and the last 1 / 3 encodes four structural proteins in turn: spike protein (S), Envelope (E), Membrane (M) and Nucleocapsid (N). Among them, the S protein contains a virus receptor binding region, which can bind to the angiotensin-converting enzyme 2 (ACE2) receptor on the surface of human cells, mediate virus adsorption and entry into cells, and is a key protein for virus invasion of host susceptible cel...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K16/10C12N15/13C12N15/85C12N5/10A61K39/42A61P31/14
CPCC07K16/10C12N15/85A61P31/14C07K2317/56C07K2317/565C07K2317/33C07K2317/76C07K2317/92A61K2039/505
Inventor 陈薇于长明李建民迟象阳孙韩聪张冠英张军范鹏飞张哲陈郑珊王步森宰晓东房婷付玲郝勐陈旖徐婧含
Owner ACADEMY OF MILITARY MEDICAL SCI
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